Fanconi Anemia (FA) is an autosomal recessive disease characterized by progressive pancytopenia, cancer susceptibility, and multiple congenital anomalies. Cells from patients with FA are hypersensitive to DNA cross- linking agents, notably mitomycin C (MMC) and diepoxybutane (DEB), and the molecular defect is presumably one of DNA repair. The cDNA for one of the known complementation groups of FA has recently been cloned yet the function for the encoded protein remains unknown. We intend to characterize differences in FACC mRNA and protein expression, transcription, replication, and DNA repair with an eventual goal of defining FACC function and lending strategies toward characterizing other FA genes. Our bank of FA cell lines includes a mutant cell line, HSC536N, that has a point mutation at one allele in the FACC gene at amino acid 554 and deletion of most of the second, that has been stably transfected and thus corrected with the wild-type Fanconi Anemia Complementation Group C (FACC) cDNA. We have used this pair of isogenic cell lines to demonstrate differences in cell cycle and cytotoxicity in response to MMC treatment and intend to further define these differences in a battery of chemotherapeutic agents, in different stages of the cell cycle by cell cycle synchronizaton studies, and in the ability to support transcription and replication using a crosslinked DNA (plasmid) template. Also, FACC mRNA and protein expression will be assayed to show differential effects from MMC treatment. Intriguingly, the FACC polypeptide appears to have some homology to DNA methyltransferase, and in vitro mutagenesis will be used to analyze this region for functional significance. In addition, the FACC gene has been localized to chromosome 9q22.3, and 20 assorted leukemia (myeloid and lymphoid) bone marrow samples have been accumulated with cytogenetically proven deletions in this region. Fluorescent in situ hybridization (FISH), DNA, and RNA analysis will be conducted on these specimens in order to demonstrate loss of heterozygosity at the FACC locus and mutations at the other allele. Other solid tumors will be similarly screened in order to test the idea that the FACC gene is a tumor suppressor or susceptibility gene.